Processes and intermediates useful to make antifolates

ABSTRACT

The present application relates to a series of novel sulfonic acid metal cation salts of the formula                    
     which are useful intermediates to prepare antifolate 5-substituted pyrrolo[2,3-d]pyrimidines. The present invention also relates to a novel process for preparing the sulfonic acid metal cation salts and to a novel process for preparing aldehydes of the formula                    
     from the corresponding sulfonic acid metal cation salts.

This application is a continuation of application Ser. No. 09/413,633filed Oct. 6, 1999 now U.S. Pat. No. 6,090,168 which is a division ofSer. No. 09/160,129 now U.S. Pat. No. 6,013,828, filed Sep. 24, 1998which claims the benefit of U.S. Provisional Application No. 60/093,039which, pursuant to 37 C.F.R. 1.53(c)(2), has a filing date of Sep. 26,1997.

FIELD OF THE INVENTION

This invention relates to synthetic organic chemistry. Specifically, theinvention relates to a process for preparing intermediates useful in thesyntheses of valuable antifolate compounds.

BACKGROUND OF THE INVENTION

Compounds known to have antifolate activity are well recognized aschemotherapeutic agents for the treatment of cancer. Recently, a seriesof 5-substituted pyrrolo[2,3-d]pyrimidine compounds of formula XVI:

where R is NHC*H(CO₂R¹)CH₂CH₂CO₂R¹ or OR¹, the configuration about thecarbon atom designated * is L, each R¹ is hydrogen or the same ordifferent carboxy protecting group, m is 2 or 3, and A is an aryl group;and their pharmaceutically acceptable salts were disclosed asantifolates or intermediates to antifolates. U.S. Pat. No. 5,416,211.

A key intermediate to compounds of formula XVI is the α-halo aldehyde offormula XV:

Among the possible routes to compounds of formula XV disclosed in U.S.Pat. No. 5,416,211, alpha halogenation of aldehydes of formula XIV:

is most direct.

A synthesis published by Taylor and Harrington teaches the route tocompounds of formula XIV shown below:

Taylor, E. C., Harrington, P. M., J.Org.Chem., 55, 3222, (1990).

Another synthesis published by Larock, et. al., may be used to form therequisite aldehydes of formula XIV by a similar palladium[0] catalyzedcoupling shown below:

Larock, R. C., Leung, W., Stolz-Dunn, S., Tet.Let., 30, 6629, (1989).

If the procedure of Larock is followed, a mixture of desired andundesired products results, the components of which are very difficultto separate and purify to afford compounds of formula XIV. In addition,regardless of how they are formed, aldehydes of formula XIV aretypically not isolated, due to their inherent instability, and areinstead alpha halogenated in situ to provide the alpha halo aldehydes offormula XIX, as disclosed in U.S. Pat. No. 5,416,211.

An improvement over the prior art would provide a facile method forselectively producing a compound of formula XIV and would provide analdehyde analogue amenable to isolation, bulk manufacturing, and storageeasily convertible to it's aldehyde form.

SUMMARY OF THE INVENTION

The present invention relates to compounds of formula IV:

where

M is a metal cation;

n is 1 or 2;

R² is NHCH(CO₂R³)CH₂CH₂CO₂R³ or OR³;

R³ is independently at each occurrence a carboxy protecting group; and

X is a bond or C₁-C₄ alk-diyl, which are useful intermediates to some ofthe antifolate 5-substituted pyrrolo[2,3-d]pyrimidines disclosed in U.S.211 which correspond to the substitution parameters of the compounds offormula IV.

The present invention further relates to a process for preparingcompounds of formula III:

where

R² is NHCH(CO₂R³)CH₂CH₂CO₂R³ or OR³; and

R³ is independently at each occurrence a carboxy protecting group;

which comprises reacting a compound of formula IV with a trialkylsilylhalide in a solvent.

The invention also relates to a process for preparing a compound offormula IV which comprises reacting a compound of formula III with acompound of the formula M(HSO₃ ⁻)_(n) in a solvent.

DETAILED DESCRIPTION OF THE INVENTION

The compounds of formula IV where R² is OR³ are enantiomeric and thecompounds of formula IV where R² is NHCH(CO₂R³)CH₂CH₂CO₂R³ arediastereomeric. Single enantiomers, single diastereomers, and mixturesthereof are encompassed within the scope of this invention. It ispreferred that the chiral center in the glutamic acid residue (R² isNHCH(CO₂R³)CH₂CH₂CO₂R³), when present, be of the “L” configuration.

In the present document, all expressions of concentration, percent,ratio and the like will be expressed in weight units unless otherwisestated, except for mixtures of solvents which will be expressed involume units. All temperatures not otherwise stated will be expressed indegrees Celsius. Compounds or compound mixtures in brackets, exceptthose brackets used to denote salt forms, signify intermediates whichare preferably not isolated before their use in subsequent reactions.

In the general formulae of the present document, the general chemicalterms have their usual meanings. For example, the term “alkyl” refers toa fully saturated, straight or branched chain, monovalent hydrocarbonylmoiety having the stated number of carbon atoms and includes, but is notlimited to, a methyl, ethyl, propyl, isopropyl, n-butyl, s-butyl, andt-butyl groups, and also includes higher homologs and isomers thereofwhere appropriate.

The term “C₁-C₄ alk-diyl” refers to a fully saturated straight chaindivalent hydrocarbon moiety having from 1 to 4 carbon atoms wherein eachcarbon atom in the chain may be independently substituted once with aC₁-C₄ alkyl group. For example, 1,2-dimethylprop-1,3-diyl is encompassedwithin the definition of C₁-C₄ alk-diyl but 1,1-dimethylprop-1,3-diyl isnot. The term is further exemplified by moieties such as, but notlimited to, —CH₂—, —CH₂CH₂—, —CH₂(CH₂)CH₂—, methyleth-1,2-diyl,—CH₂(CH₂)₂CH₂—, and but-1,3-diyl. Preferred C₁-C₄ alk-diyl groups arethose that are unsubstituted and most preferred are —CH₂— and —CH₂CH₂—.

The term “C₂-C₆ alkenyl” refers to a mono-unsaturated, monovalent,hydrocarbon moiety containing from 2 to 6 carbon atoms which may be in abranched or straight chain configuration. The term is exemplified bymoieties such as, but not limited to, ethylenyl, propylenyl, allyl,butylenyl, and pentylenyl.

The term “C₁-C₄ alkoxy” refers to a methoxy, ethoxy, propoxy,isopropoxy, butoxy, s-butoxy, and a t-butoxy group.

The term “halo” and “halide” refers to chloride, bromide, or iodide.

The terms “substituted benzyl”, “substituted benzhydryl”, and“substituted trityl” refers to a benzyl, benzhydryl, and trityl group,respectively, substituted from 1 to 5 times independently with a nitro,C₁-C₄ alkoxy, C₁-C₆ alkyl, or a hydroxy(C₁-C₆ alkyl) group. Thesesubstitutions will only occur in a sterically feasible manner such thatthe moiety is chemically stable.

The terms “substituted C₁-C₆ alkyl” and “substituted C₂-C₆ alkenyl”refer to a C₁-C₆ alkyl and C₂-C₆ alkenyl group respectively substitutedfrom 1 to 3 times independently with a halo, phenyl, tri(C₁-C₄alkyl)silyl, or a substituted phenylsulfonyl group.

The terms “substituted phenyl” and “substituted phenylsulfonyl” refer toa phenyl and phenylsulfonyl group respectively where the phenyl moietyof either is para substituted with a C₁-C₆ alkyl, nitro, or a halogroup.

The term “leaving group” refers to a monovalent substituent of amolecule which is prone to nucleophilic displacement. Typical leavinggroups include, but are not limited to, sulfonates such as phenyl,substituted phenyl, C₁-C₆ alkyl, and C₁-C₆ perfluoro alkylsulfonates;halides; and diazonium salts such as diazonium halides.

The term “carboxy protecting group” as used in this specificationdenotes groups which generally are not found in the final therapeuticcompounds but are intentionally introduced during a portion of thesynthetic process to protect a group which otherwise might react in thecourse of chemical manipulations, and is later removed. Examples of suchcarboxylic acid protecting groups include C₁-C₆ alkyl, substituted C₁-C₆alkyl, C₂-C₆ alkenyl, substituted C₂-C₆ alkenyl, benzyl, substitutedbenzyl, benzhydryl, substituted benzhydryl, trityl, substituted trityl,trialkylsilyl, aroyl groups such as phenacyl, and like moieties. Thespecies of carboxy-protecting group employed is not critical so long asthe derivitized carboxylic acid is stable to the conditions ofsubsequent reaction(s) on other positions of the molecule and can beremoved at the appropriate point without disrupting the remainder of themolecule. Carboxy protecting groups similar to those used in thecephalosporin, penicillin, and peptide arts can also be used to protecta carboxy group substituent of the compounds provided herein. Futherexamples of these groups are found in E.Haslam, “Protective Groups inOrganic Chemistry”, J. G. W. McOmie, Ed., Plenum Press, New York, N.Y.,1981, Chapter 5 and T. W. Greene, “Protective Groups in OrganicSynthesis”, 2nd Ed., John Wiley and Sons, New York, N.Y., 1991, Chapter5. When R¹ or R³ is a carboxy protecting group, the protecting group ispreferably C₁-C₄ alkyl. The most preferred protecting groups are methyland ethyl.

The term “trialkylsilyl” refers to a monovalent silyl group substituted3 times independently with a C₁-C₆ alkyl group.

The term “trialkylsilyl halide” refers to a compound of the formula(C₁-C₆ alkyl)₃—Si-halo wherein each C₁-C₆ alkyl is the same ordifferent. Trialkylsilyl halides include, but are not limited to,trimethylsilyl, triethylsilyl, tripropylsilyl chloride, bromide, andiodide.

The term “metal cation” refers to an alkali or alkaline earth metalcation. Alkali metals form singly charged cations e.g., Li⁺¹, Na⁺¹, andK⁺¹, while alkaline earth metals form doubly charged cations e.g., Mg⁺²and Ca⁺² but the charge on the compounds of formula IV, on compounds ofthe formula M(HSO₃ ⁻)_(n), or on metal cation chlorides as a whole, iszero. Therefore, when M is a Group I metal, the molar ratio betweencation and anion is 1:1 and when M is a Group II metal cation, the molarratio is 1:2.

The term “pharmaceutical salt” as used herein, refers to salts preparedby reaction of the compounds of the present invention with a mineral ororganic acid (e.g. hydrochloric, hydrobromic, hydroiodic, orp-toluenesulfonic acid) or an inorganic base (e.g. sodium, potassium,lithium, magnesium, or hydroxide, carbonate, or bicarbonate). Such saltsare known as acid addition and base addition salts. For furtherexemplification and methods of preparing pharmaceutical salts, see e.g.Berge, S. M, Bighley, L. D., and Monkhouse, D. C., J. Pharm. Sci., 66,1, 1977.

The term “phase transfer catalyst” refers to a salt in which the cationhas large nonpolar substituent groups which confer good solubility onthe salt in organic solvents. The most common examples aretetraalkylammonium and tetraalkylphosphonium ions e.g.tetraalkylammonium chloride or bromide.

The term “palladium catalyst” refers to a reagent which is a source ofpalladium zero (Pd(0)). Suitable sources of Pd(0) include, but are notlimited to palladium(0) bis(dibenzylidineacetone) and palladium(II)acetate.

The term “halogenating reagent” refers to a reagent that can provide anelectrophilic source of a halogen to the target molecule. Typicalhalogenating reagents include but are not limited to benzeneseleninylchloride, bromide, or iodide, thionyl bromide or chloride,dibromobarbituric acid, N-bromo-, N-iodo-, and N-chloro succinimide,elemental chlorine, elemental bromine (and complexes of bromine such asbromine dioxane complex), and elemental iodine, and the like.

The term “thermodynamic base” refers to a base which provides areversible deprotonation of an acidic substrate or is a proton trap forthose protons that may be produced as byproducts of a given reaction,and is reactive enough to effect the desired reaction withoutsignificantly effecting any undesired reactions. Examples ofthermodynamic bases include, but are not limited to, acetates, acetatedihydrates, carbonates, bicarbonates, and hydroxides (e.g. lithium,sodium, or potassium acetate, acetate dihydrate, carbonate, bicarbonate,or hydroxide), tri-(C₁-C₄ alkyl)amines, or aromatic nitrogen containingheterocycles (e.g. imidazole and pyridine).

The term “suitable solvent” refers to any solvent, or mixture ofsolvents, inert to the ongoing reaction that sufficiently solubilizesthe reactants to afford a medium within which to effect the desiredreaction.

Synthesis

Compounds of formula IV may be prepared by a novel process illustratedin Scheme 1 below where Lg is a leaving group, R⁴ is hydrogen or C₁-C₄alkyl, and X′ is C₁-C₄ alk-diyl;

with the proviso that if X′ is not a bond, then the

carbon alpha to the alcohol must be a —CH₂— moiety; and n, R² and X areas defined above for formula IV.

A mixture containing a compound of formula III may be prepared bydissolving or suspending a compound of formula II in a suitable solvent,in the presence of a suitable thermodynamic base and a phase transfercatalyst, optionally in the presence of a metal cation chloride, andadding a compound of formula I and a palladium catalyst. Once all thereactants are combined, the reaction may be conducted at temperaturesranging from at least about 0° C. to about 100° C. Within this broadtemperature range, when Lg is bromide in compounds of formula II, thereaction mixture should be heated to at least about 50° C., morepreferably at least about 60° C., and most preferably at least about 65°C. for from about 8 to about 24 hours. When Lg is iodide, the reactionproceeds more robustly, thus a temperature range of 0° C. to about 25°C. is the typical temperature range with room temperature being thepreferred reaction temperature. The reaction is preferably allowed torun for from 8 to about 10 hours.

Suitable solvents for this reaction include, but are not limited to,dimethylsulfoxide, tetrahydrofuran, N,N′-dimethylimidazole, diethylether, dimethoxyethane, dioxane, acetonitrile, mixtures thereof, and thelike. Typically, an alkali metal acetate is generally the preferredthermodynamic base, and lithium acetate is the particularly preferredbase. However, when Lg is bromo, lithium acetate dihydrate is thepreferred base. In general, dimethylformamide or dimethylacetamide isthe preferred solvent. Tetrabutylammonium bromide is generally thepreferred phase transfer catalyst. Palladium(II) acetate is typicallythe preferred palladium catalyst. Although not required, it is preferredto employ an alkali metal chloride in order to maximize the yield of thedesired product of formula III. Lithium chloride is the preferred metalcation chloride. Preferred compounds of formula I are those where R⁴ ishydrogen and X′ is a bond, —CH₂—, or —CH₂CH₂—. In compounds of formulaII, Lg is preferably bromo, iodo, or trifluoromethylsulfonyloxy. Themost preferred Lg moiety is iodo. The most preferred compound of formulaI is 3-butenol.

Relative to the compounds of formula II, the following amounts ofpreferred reagents are typically employed:

thermodynamic base—1.0 to about 3.0, preferably about 1.05 to about 1.3equivalents;

metal cation chloride—0 to about 4, preferably about 2.8 to about 3.2equivalents;

phase transfer catalyst—0 to about 3.0, preferably 0.4 to about 0.6equivalents; and

palladium catalyst—0.015 to about 0.1, preferably about 0.02 to about0.03 equivalents.

compound of formula I—1.0 to about 2.0, preferably about 1.1 to about1.3 equivalents.

The reaction discussed above results in a mixture of products whichincludes a compound of formula III, which may be isolated but ispreferably further reacted as described in Scheme 1. Substantialpurification of the compound of formula III or separation from theundesired byproducts is not necessary before proceeding to the nextnovel step in the overall process. Preferably, a simple extraction usingan aqueous immiscible solvent followed by filtration of the palladiumcatalyst is all that is performed before proceeding. Suitable solventsfor the extraction include, but are not limited to, methylene chloride,chloroform, methyl acetate, carbon tetrachloride, mixtures thereof, andthe like. The preferred solvent is typically ethyl acetate.

A metal bisulfite reactant of the formula M(HSO₃ ⁻)_(n) may be added tothe organic extract filtrate from above (the mixture that contains acompound of formula III and byproducts). Typically, a lower alcohol,preferably ethanol 5% denatured with methanol (3A ethanol) or ethanol0.5% denatured with toluene (2B-3 alcohol), and water are also added asco-solvents for this reaction. The volume of ethanol added is preferablyabout equal to that of the ethyl acetate originally present while thevolume of water in the mixture is proportional to the volume ofdenatured ethanol, preferably at a ratio of about 1:5. Suitable metalbisulfite reactants include, but are not limited to, sodium bisulfite(NaHSO₃), potassium bisulfite (KHSO₃), lithium bisulfite (LiHSO₃) andmagnesium bisulfite (Mg(HSO₃)₂). A preferred metal bisulfite reactant issodium bisulfite. The amount of metal bisulfite reactant employedtypically ranges from about 0.85 equivalents to about 1.2 equivalents,relative to the compound of formula III. The preferred amount of metalbisulfite reactant is typically about 0.90 to 1.1 and is most preferablyabout 0.95 to 1.0 equivalents. The reaction may be performed for from 2to about 15 hours at a temperature range from room temperature to about55° C. It is preferred to conduct the reaction for a time of betweenabout 2 and 5 hours at a temperature of between about 35° C. and about50° C.

When the reaction is complete, different amounts of various sulfonicacid metal cation salt products are created depending on the makeup ofthe mixture which contained the compound of formula III. The majorcomponent is the sulfonic acid metal cation salt of formula IV.Typically, the major component compound of formula IV will precipitateout of the product mixture spontaneously, but where spontaneouscrystallization does not occur, it is possible by careful adjustment ofthe solvent volumes to cause the major component to crystallize.Usually, the amount of ethyl acetate relative to both the ethanol andwater is increased in order to force the precipitation of the majorcomponent sulfonic acid metal cation salt. This technique of adjustmentof solvent volumes is well known to those skilled in the art. Onceprecipitated, the desired major component sulfonic acid metal cationsalt of formula IV may then be collected via filtration.

The preferred compounds of formula IV are:

Application of the above chemistry enables the synthesis of thecompounds of formula IV, which include, but are not limited to:

1-hydroxy-3-(4-carbomethoxyphenyl)propanesulfonic acid sodium salt;

1-hydroxy-3-(4-carboethoxyphenyl)propanesulfonic acid potassium salt;

1-hydroxy-2,3-dimethyl-4-(4-carbomethoxyphenyl)butanesulfonic acidlithium salt;

N-(4-[(3-hydroxy-3-sulfonic acid sodium salt)propyl]benzoyl)-L-glutamicacid dimethyl ester;

N-(4-[(3-hydroxy-3-sulfonic acid potassiumsalt)propyl]benzoyl)-L-glutamic acid diethyl ester;

N-(4-[(1,2-dimethyl-4-hydroxy-4-sulfonic acid lithiumsalt)butyl]benzoyl)-L-glutamic acid dipropyl ester;

Compounds of formula III may be prepared from compounds of formula IV bya novel process shown in Scheme 2 below where M, n, R², and X are asdefined above for formula IV.

Compounds of formula IV can be converted to aldehydes of formula III bydissolving or suspending a compound of formula IV in a suitable solventand adding a trialkylsilyl halide. Suitable solvents include, but arenot limited to, acetone, tetrahydrofuran, diethylether, methylenechloride, methyl acetate, ethyl acetate, chloroform, mixtures thereof,and the like. The preferred solvent is typically acetonitrile. It hasbeen found that yields for this reaction can be increased by degassingthe solution containing the compound of formula IV, before the additionof the trialkylsilyl chloride, with an inert gas. Typically, nitrogen isemployed as the inert gas. The preferred trialkylsilyl halide is usuallytrimethylsilyl chloride. The trialkylsilyl halide is typically employedin a stoichiometric excess. For example, a 2 to 4 stoichiometric excess,relative to the compound of formula IV is typically employed. A 2.7 toabout 2.9 stoichiometric excess is usually preferred. The mixture istypically allowed to react for from about fifteen minutes to about onehour. The reaction is usually performed at an elevated temperature of atleast about 30° C., preferably at least about 40° C., more preferably atleast about 50° C., and most preferably the mixture is allowed to run atbetween about 60° C. and 70° C.

Although isolation and purification of the compounds of formula IIIformed by the overall novel process of this invention is possible, thesecompounds are typically not substantially purified but are insteadconverted to 5-substituted pyrrolo[2,3-d]pyrimidine compounds of formulaVII(a) by the process shown in Scheme 3 below where R² and X are asdefined above for formula IV.

Compounds of formula V may be prepared by adding a halogenating reagentto the solution containing the compound of formula III prepared asdescribed in Scheme 2. The addition may occur at the preferred 60° C. to70° C. reaction temperature of the previous reaction but the reaction ispreferably cooled before the addition of the halogenating reagent. Theaddition of the halogenating reagent may be done at a temperature offrom 0° C. to 60° C., but it has been found that an addition temperatureof about 35° C. to about 45° C. is preferred. Once the halogenatingagent is added, the resulting mixture is stirred for from about 5minutes to about 2 hours. In general, time for the halogenation reactionis from about 5 minutes to about 1 hour, but is preferably performed in20 minutes or less. The preferred halo substituent in compounds offormula V is bromo and the preferred halogenating agent is typicallyelemental bromine. Once the reaction is complete, it may be quenched bythe addition of an aqueous solution of a known halogen scavenger such assodium bisulfite. The compound of formula V may then be extracted into asuitable, aqueous immiscible organic solvent. This solution whichcontains the compound of formula V is of high purity and may be useddirectly to prepare compounds of formula VII(a) or compounds of formulaVII:

and their pharmaceutical salts and solvates; by following the proceduresdescribed in U.S. Pat. No. 5,416,211, the teachings of which are hereinincorporated by reference.

When any of the compounds of formula II, IV, IV, VII, or VII(a) containcarboxy protecting groups, they may be removed by well known methods inthe art. Numerous reactions for the installation and removal of thecarboxy protecting groups contemplated within the scope of thisinvention are described in a number of standard works including, forexample The Peptides, Vol. I, Schrooder and Lubke, Academic Press(London and New York, 1965) and the Greene reference cited above.Methods for removing preferred carboxy protecting groups, particularlymethyl and ethyl groups, are essentially as described in Examples 5 and7 infra.

When R is NHCH(CO₂R¹)CH₂CH₂CO₂R¹ in compounds of formula VII or when R²is NHCH(CO₂R³)CH₂CH₂CO₂R³ in compounds of formula II, IV, IV, or VII(a),the R or R² group can be installed at any convenient point in thesynthesis. For example, the glutamic acid residue may be installed afterthe reactions of Schemes 1-3 essentially as described in Examples 5 and6 infra. In the alternative, a commercially available glutamic aciddialkyl ester of the formula NH₂CH(CO₂R³)CH₂CH₂CO₂R³ may be coupled witha commercially available p-halobenzoic acid before subsequent reactionin Scheme 1.

The optimal time for performing the reactions of Schemes 1-3 can bedetermined by monitoring the progress of the reaction by conventionalchromatographic techniques. Choice of reaction solvent is generally notcritical so long as the solvent employed is inert to the ongoingreaction and sufficiently solubilizes the reactants to afford a mediumwithin which to effect the desired reaction. Unless otherwise indicated,all of the reactions described herein are preferably conducted under aninert atmosphere. The preferred inert atmosphere is nitrogen.

Advantages/Distinctions Over the Prior Art

The process illustrated in Scheme 1 for preparing the novel compounds offormula IV greatly simplifies the purification of compounds of formulaIII formed by the alkenol coupling to an aryl halide. The processillustrated in Scheme 2 is a previously unknown method of generatingaldehydes from sulfonic acid metal cation salts. That conversion isexpected to be generally applicable and has great potential for generalsynthetic utility. Specific to this case, the conversion generatesselectively and cleanly the compounds of formula III. In addition, thecompounds of formula IV, which can be considered aldehyde analogues inthe context of this invention, are stable, usually crystalline materialsamenable to bulk manufacture, purification, and storage. Thus, ingeneral, commercial processes which require aldehydes of the formulaIII, or similar aldehydes, are made simpler by the overall process ofthe present invention.

The following examples are illustrative only and are not intended tolimit the scope of the invention in any way. The terms and abbreviationsused in the instant examples have their normal meanings unless otherwisedesignated. For example “° C.”, “N”, “mmol”, “g”, “d”, “mL”, “M”,“HPLC”, “¹H-NMR”, “¹³C-NMR”, and “vol.” refers to degrees Celsius,normal or normality, millimole or millimoles, gram or grams, density,milliliter or milliliters, molar or molarity, high performance liquidchromatography, proton nuclear magnetic resonance, carbon-13 nuclearmagnetic resonance, and an amount in mL/grams relative to startingmaterial respectively. In addition, the absorption maxima listed for theIR spectra are only those of interest and not all of the maximaobserved.

EXAMPLES Example 1 4-(4-Carbomethoxyphenyl)butanal

The Deloxan® THP Type 2 Resin used below was pretreated by mixing itwith isopropyl alcohol (2.0 vol. 20 mL) and washing with ethyl acetate(4.0 vol., 40 mL). The organic layer/resin slurry was then filteredbefore subsequent use as described below.

4-Bromobenzoic acid, methyl ester (60.0 g, 279.00 mmol), lithium acetatedihydrate (31.31 g, 306.90 mmol), lithium chloride (35.48 g, 837 mmol),and tetrabutylammonium chloride (41.22 grams, 131.49 mmol) were added todimethylformamide (698 mL). The resulting solution was degassed with asubsurface nitrogen purge. 3-buten-1-ol (24.19 grams, 28.81 mL, 334.81mmol) and palladium acetate (1.57 grams, 6.98 mmol) were added and thereaction mixture was heated to 65° C. with stirring for approximately 10hours. Reaction completion was indicated by starting materialconsumption (less than 0.4% 4-bromobenzoic acid, methyl ester remaining)as shown by HPLC (reverse phase, 60% acetonitrile:2.5% acetic acidbuffer). The reaction mixture was cooled to 25° C.-30° C. and water (700ml) and ethyl acetate (700 mL) were added. The reaction mixture wasstirred for 10 minutes and subsequently the layers were allowed toseparate. The organic layer was separated and retained and the aqueouslayer was extracted two additional times with ethyl acetate (720 mL).The ethyl acetate washes were combined with the original organic layerand the combined organic layers were washed with brine (350 mL). Theorganic layer was filtered, to remove elemental palladium, and slurriedwith Deloxan® THP Type II Resin (3.0 grams dry weight) for 45 minutes.The title compound was obtained as a solution in ethyl acetate, inapproximately 87% yield. A small amount of ethyl acetate solution wasconcentrated for characterization of product.

Analytical Data:

¹H NMR:(d₆-DMSO) δ 9.65 (t, J=1.5 Hz, 1H), 7.86 (d, J=8.5 Hz, 2H), 7.32(d, J=8.5 Hz, 2H), 3.82 (s, 3H), 2.63 (t, J=7.7 Hz, 2H), 2.43 (td,J=7.4, 1.5 Hz, 2H), 1.82 (m, 2H). ¹³C-NMR: (d₆-DMSO) δ 203.1, 166.2,147.4, 129.3, 128.7, 127.4, 51.9, 42.4, 34.3, 23.0.

Example 2 1-Hydroxy-4-(4-Carbomethoxyphenyl)butanesulfonic Acid SodiumSalt

The ethyl acetate extracts from Example 1 were concentrated to 3.6 vol.(8.7 mL) in vacuo at about 37° C. 3A Alcohol (3 vol., 7.2 mL) and water(0.63 vol., 1.51 mL) were added followed by sodium bisulfite (1.04 g,10.03 mmol). The reaction mixture was stirred for approximately 8 hours.After 10 minutes crystallization of the sulfonic acid began. Reactioncompletion was determined by ¹H NMR analysis of the reaction mixturefiltrate. The resulting white slurry is filtered to afford the titlecompound (2.78 grams, 8.98 mmol) as a white crystalline solid inapproximately 80% yield. The filter cake was washed with ethanol (1.8vol.) and dried in vacuo at 40° C. Isomeric impurities werenon-detectable by NMR.

Analytical Data:

¹H-NMR: (d₆-DMSO) δ 7.86 (d, J=8.27 Hz, 2H), 7.32 (d, J=8.27 Hz, 2H),5.33 (d, J=2.3 Hz, 1H), 3.84 (m, 1H), 3.81 (s, 3H), 2.63 (m, 2H), 1.75(m, 1H), 1.73 (m, 1H), 1.61 (m, 1H), 1.48 (m, 1H). ¹³C-NMR: (d₆-DMSO) δ166.2, 148.3, 129.2, 128.7, 127.1, 82.7, 51.9, 35.1, 31.2, 27.2. IR:(run as KBr pellet) 3237, 2962, 2930, 2889, 1726 cm⁻¹.

Example 3 1-Hydroxy-2-Bromo-4-(4-Carbomethoxyphenyl)butanal

To a 50 mL round bottom flask with magnetic stirrer were added4-(4-oxobutyl)-benzoic acid methyl ester sodium bisulfite adduct (3.10grams, 10 mmol), acetonitrile (14 mL) and chlorotrimethylsilane (3.6 mL,28 mmol). Nitrogen gas was bubbled through for five minutes and then themixture was heated in a 60° C. bath for one hour under nitrogen. Themixture at this point in time was a light yellow. The mixture was thencooled under refrigeration to 5° C. and bromine (0.5 mL, 9.7 mmol,) wasadded. The brownish bromine color was discharged within 1 minute. Thesolution was light yellow and the visible solids appeared colorless. Themixture was removed from the cooling bath and stirred for an additional2 hours. Water (14 mL) and sodium bisulfite (0.14 grams) were added toscavenge/quench the remaining bromine and the resulting mixture stirredfor 1 hour. The mixture was then partitioned between methylene chloride(14 mL) and an additional 7 mL of water. The organic phase was separatedand stripped on a rotary evaporator until only 26 mL remained. Withinthis 26 mL is the title compound which was not purified or isolatedfurther before subsequent reaction as in Example 4 below. A small amountof the methylene chloride solution was concentrated for characterizationof product.

Analytical Data:

¹H-NMR: (CDCl₃) δ 9.40 (d, 1H), 7.95 (d, 2H), 7.26 (d, 2H), 4.15 (ddd,1H), 3.88 (s, 3H), 2.89 (m, 1H), 2.79 (m, 1H), 2.35 (m, 1H), 2.21 (m,1H). ¹³C-NMR: (CDCl₃) δ 191.4, 166.8, 145.1, 129.9, 128.5, 128.5, 54.4,52.0, 32.7, 32.6.

Example 44-[2-(2-Amino-4,7-Dihydro-4-Oxo-1H-Pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoicAcid Methyl Ester

The 26 mL of organic layer from Example 3, which contains1-hydroxy-2-bromo-3-(4-carbomethoxyphenyl)butanal, had added to it2,4-diamino-6-hydroxy pyrimidine (1.26 grams, 10 mmol), sodium acetate(1.68 grams, 20 mmol) and water (23 mL). Nitrogen was bubbled throughthis reaction mixture for 5 minutes. The mixture was heated at 40° C.under N₂ for 2 hours. The mixture was cooled to ambient conditions andfiltered and the collected solids were washed with 23 mL of a 1:1mixture of acetonitrile and water. The filter cake was dried to yield1.47 grams of light yellow needles. The analysis showed a 45% overallyield for Examples 3 and 4 and also showed the title compound wasproduced at a purity level of 94.8% by HPLC (reverse phase, gradient 50%to 30% methanol:20 mM potassium dihydrogen phosphate or ammoniumdihydrogen phosphate buffer).

Analytical Data:

¹H-NMR (d6-DMSO) δ 10.66 (s, 1H), 10.23 (s, 1H), 7.84 (d, 2H), 7.32 (d,2H), 6.31 (s, 1H), 6.08 (s, 2H), 3.80 (s, 3H), 2.98 (dd, 2H), 2.86 (dd,2H). ¹³C-NMR (d₆-DMSO) δ 166.3, 159.4, 152.3, 151.3, 148.4, 129.2,128.7, 127.1, 117.6, 113.6, 98.8, 52.0, 36.3, 27.9.

Example 54-[2-(2-Amino-4,7-Dihydro-4-Oxo-1H-Pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoicAcid

A flask was charged with 13.0 grams of4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoicacid, methyl ester and 150 mL of 2N aqueous sodium hydroxide solution.Stirring was applied and the slurry was heated to 40° C. The reactionwas monitored by HPLC (reverse phase, gradient 50% to 30% methanol:20 mMpotassium dihydrogen phosphate or ammonium dihydrogen phosphate buffer).3A Alcohol (230 mL) was added to the solution, which was then seededwith authentic4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoicacid (obtained by following the procedure of U.S. Pat. No. 5,416,211).The solution pH was adjusted to 4.4 with 6N hydrochloric acid (48.5 mL).The solids were filtered off and washed with 30 mL of a 1:1 mixture ofwater:3A alcohol. The solids were dried in vacuo at 50° C. 10.84 gramsof the title compound were recovered.

Analytical Data:

¹H NMR (d6-DMSO) δ 10.66 ′(br s, 1H), 10.33 (br s, 1H), 7.83 (d, 2H),7.30 (d, 2H), 6.31 (s, 1H), 6.17 (br s, 2H), 2.97 (m, 2H), 2.85 (m, 2H).¹³C-NMR (d₆-DMSO) δ 167.6, 159.5, 152.4, 151.4, 147.9, 129.4, 128.6,128.4, 117.7, 113.6, 98.8, 36.4, 28.0.

Example 6N-(4-[2-(2-Amino-4,7-Dihydro-4-Oxo-1H-Pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl)-L-GlutamicAcid Diethyl Ester p-Toluenesulfonic Acid Salt

A 50 mL flask with mechanical stirrer, thermometer and N₂ adapter wascharged with 1.93 g (corrected for assay) of4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoicacid (2.5 g, potency 77%) and 13.5 mL of dimethylformamide. The slurrywas stirred 15 minutes and 1.94 grams of N-methylmorpholine (2.9 eq) wasadded. The mixture was cooled to 5° C. with an ice/water bath andchlorodimethoxytriazine (1.46 grams, 1.28 eq.) was added all at once.The mixture was stirred 40 minutes before L-glutamic acid diethyl ester(1.99 g, 1.28 eq) was added all at once. The reaction was allowed towarm to ambient temperature. The reaction was monitored by HPLC (reversephase, gradient 20% to 46% acetonitrile:0.5% acetic acid buffer) and wascomplete in 1 hour at 23° C. The reaction mixture was transferred to a250 mL Erlenmeyer flask containing 36 mL of deionized water and 18 mL ofmethylene chloride. The reaction flask was rinsed with 18 mL ofmethylene chloride which was added to the Erlenmeyer flask. The mixturewas stirred 15 minutes and the layers were allowed to separate. Themethylene chloride layer was concentrated from 46 grams to 13 gramsusing a rotary evaporator at reduced pressure at a bath temperature at45° C. The concentrate was diluted with 55 mL of 3A alcohol, andconcentrated again to 10 grams to remove methylene chloride. Theconcentrate was diluted to a total volume of 60 mL with 3A alcohol andthe resulting solution was heated to 70° C. to 75° C. p-Toluenesulfonicacid (3.16 g, 2.57 eq.) dissolved in 55 mL of 3A alcohol were added over30-90 minutes. The resulting slurry was refluxed for an hour. The slurrywas cooled to ambient temperature and filtered using a 7 cm Buchnerfunnel. The wet cake was washed with 25 mL ethanol and dried in vacuo at50° C. overnight to yield 3.66 grams of the title compound. Potency 95%

Analytical data:

¹H NMR (d₆ DMSO) δ 11.59 (br s, 1H), 11.40 (s, 1H), 8.66 (d,1H), 7.88(brs, 1H), 7.79 (d, 2H), 7.58 (d, 2H), 7.29 (d, 2H), 7.16 (d, 2H), 6.52 (s,1H), 4.42 (m, 1H), 4.09 (q, 2H), 4.03 (q, 2H), 2.94 (m, 2H), 2.89 (m,2H), 2.43 (m, 2H), 2.28 (s, 3H), 2.08 (m, 1H), 2.02 (m, 1H), 1.17 (t,3H), 1.14 (t, 3H). ¹³C NMR (d₆ DMSO) δ 172.3, 171.9, 166.7, 157.2,150.6, 145.8, 144.4, 138.6, 138.3, 131.3, 128.4, 128.3, 127.5, 125.6,119.2, 115.4, 99.1, 60.6, 60.0, 52.0, 35.8, 30.2, 27.2, 25.8, 20.8,14.1, 14.1.

Example 7N-[4-[2-(2-Amino-4,7-Dihydro-4-Oxo-1H-Pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-GlutamicAcid

To 1.00 gram ofN-[4-[2-(2-amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamicacid, diethyl ester p-toluenesulfonic acid salt in a 50 ml Erlenmeyerflask was added 1N aqueous sodium hydroxide (6.7 mL) and the resultingmixture stirred until all the solids had dissolved (approximately 20minutes). The solution was light green. An additional 6-7 mL ofdeionized water was added and the pH was adjusted to 2.8-3.1 with dilutehydrochloric acid. The resulting slurry was heated to approximately 70°C. in order to produce larger particles of solids. The solids werefiltered to yield the title compound.

Example 8N-(4-[2-(2-Amino-4,7-Dihydro-4-Oxo-1H-Pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl)-L-GlutamicAcid Disodium Salt

N-[4-[2-(2-Amino-4,7-dihydro-4-oxo-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]-L-glutamicacid from Example 7 was dissolved in 3.8 mL of water and 2.2 ml of 1Nsodium hydroxide. The pH of the mixture was adjusted to 7.5-8.5 usingdilute hydrochloric acid and 1N sodium hydroxide. The solution washeated to 70° C. and 40 mL of 3A alcohol were added. The solution wasallowed to cool to room temperature during which time a thick slurrydeveloped. The solids were filtered and washed with 4:1 ethanol:water.The solids were dried at 50° C. in a vacuum oven. 640 milligrams of thetitle compound were recovered as a solid.

Analytical Data:

¹H NMR (300 MHz, DMSO-d₆/D₂O) δ 7.67 (d, J=7.8 Hz, 2H), 7.22 (d, J=7.8Hz, 2H), 6.30 (s, 1H), 4.09 (m, 1H), 2.88 (m, 2H), 2.83 (m, 2H),2.05-1.71 (m, 4H). ¹³C NMR (75 MHz, DMSO-d₆/D₂O) δ 179.9, 176.9, 167.1,160.8 152.9, 151.7, 146.7, 132.6, 129.4, 127.9, 118.7, 115.2, 99.5,56.1, 36.8, 35.3, 30.1, 28.4.

Example 91-Hydroxy-4-(L-N-[1,3-Dicarboethoxypropyl]benz-4-amide)butanesulfonicAcid Sodium Salt

L-N-1,3-(Dicarboethoxypropyl)-4-iodobenzamide (10.00 g, 23.1 mmol),lithium chloride (2.937 g, 69.3 mmol), lithium acetate (2.592 g, 25.4mmol), tetrabutylammonium chloride (3.412 g, 11.55 mmol) anddimethylformamide (57.7 mL) were combined. The mixture was thoroughlysparged with nitrogen. 3-buten-1-ol (1.998 g, 27.7 mmol) andpalladium(II)acetate (0.130 g, 0.577 mmol) were added. The mixture washeated to 60° C. under nitrogen for 24 hours. At this point HPLC(reverse phase, 60% acetonitrile:2.5% acetic acid buffer) indicatedreaction completion. The reaction was partioned between ethyl acetate(58 mL) and water (58 mL). The layers were separated. The aqueous layerwas extracted twice with ethyl acetate (58 mL per extraction). Theorganic layers were combined and washed with brine (30 mL). Theresulting organic layer was concentrated to 25 mL. Ethyl acetate (15mL), water (3.25 mL), and sodium bisulfite (0.636 g, 6.11 mmol) wereadded. The mixture was stirred at 25° C. for 16 hours. Acetone (75 mL)was added. The product precipitate was collected by filtration and driedin a vacuum oven to give 1.59 g of the title compound. Yield: 48.6%.

Example 10N-(4-[2-(2-Amino-4,7-Dihydro-4-Oxo-1H-Pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl)-L-GlutamicAcid Diethyl Ester

In a 25 mL round bottom flask with magnetic stirring were combined1-hydroxy-4-(L-N-[1,3-dicarboethoxypropyl]benz-4-amide)butanesulfonicacid sodium salt (0.922 g, 2.0 mmol), acetonitrile (5 mL) andtrimethylsilyl chloride (0.72 mL). The mixture was sparged with nitrogenfor 5 minutes, and then heated to 60° C. for 1 hour. The temperature wasadjusted to 40° C. and bromine (98 μL, 1.9 mmol) was added. ¹H-NMRindicated clean conversion to the α-bromide intermediate. The reactionwas cooled to ambient and washed with 1% aqueous sodium bisulfitesolution (2.5 mL). The organic phase was stripped to an oil.2,4-diamino-6-hydroxypyrimidine (300 mg, 2.4 mmol), sodium acetate (500mg), water (5 mL), and acetonitrile (5 mL) were added. The mixture washeated at 40° C. for 6 hours. The upper, organic phase was collected andconcentrated to an oil (450 mg). ¹H-NMR and HPLC (reverse phase,gradient 20% to 46% acetonitrile:0.5% acetic acid buffer) confirmed thatthe oil was predominately the title compound.

The present invention has been described in detail, including thepreferred embodiments thereof. However, it will be appreciated thatthose skilled in the art, upon consideration of the present disclosure,may make modifications and/or improvements that fall within the scopeand spirit of the invention as set forth in the following claims.

I claim:
 1. A process for preparing a compound of the formula:

wherein: R³′ is hydrogen or a carboxy protecting group; and X is a bondor C₁-C₄ alk-diyl; which comprises: a) reacting a compound of theformula:

 wherein: R³ is a carboxy protecting group; M is a metal cation; and nis 1 or 2; with a trialkylsilyl halide in a solvent; b) adding ahalogenating reagent to the reaction solution of step a) to form acompound of the formula:

 wherein halo is chloride, bromide or iodide; c) reacting the product ofstep b) with a compound of formula VI:

 in a solvent; and d) optionally removing the carboxy protecting groupfrom the product of step c).
 2. The process according to claim 1 whereinhalo is bromide; R³ is C₁-C₄ alkyl; X is a bond, —CH₂— or —CH₂CH₂—; andthe trialkylsilyl halide is trimethylsilyl chloride.
 3. The processaccording to claim 2 wherein the halogenating reagent is elementalbromine; n is 1; R³ is methyl; X is —CH₂—; the alkali metal is sodium;and the step a) solvent is acetonitrile.
 4. The process according toclaim 3 wherein the step a) reaction is performed at a temperaturebetween 50° C. and 70° C. and the step b) reaction is performed between35° C. and 45° C.
 5. The process according to claim 1 wherein step d) isperformed and which further comprises: a) installing a L-glutamic acidresidue of the formula NHC*H(CO₂R³)CH₂CH₂CO₂R³ to prepare a compound ofthe formula

 or a salt thereof; wherein the configuration about the carbon atomdesignated * is L and R³ is independently at each occurrence a carboxyprotecting group; b) optionally removing the carboxy protecting groupsfrom the product of step a); and c) optionally salifying the product ofstep a) or step b) to form a compound of the formula:

 or a salt thereof.
 6. The process according to claim 5 wherein X is abond, —CH₂— or —CH₂CH₂—; R³ is independently C₁-C₄ alkyl at eachoccurrence; and the trialkylsilyl halide is trimethylsilyl chloride. 7.The process according to claim 6 wherein n is 1; R³ is ethyl at eachoccurrence within the L-glutamic acid residue but is methyl within theclaim 1, step a) starting material; X is —CH₂—; the alkali metal issodium; and the claim 1 step a) solvent is acetonitrile.
 8. The processaccording to claim 7 wherein the claim 1 step a) reaction is performedat a temperature between 50° C. and 70° C. and the claim 1 step b)reaction is performed between 35° C. and 45° C.
 9. The process accordingto claim 8 wherein said salt is the sodium hydroxide base addition salt.10. The process according to claim 9 wherein said sodium hydroxide baseaddition salt is the compound of the formula: